Tularemia is the severe and sometimes fatal disease caused by infection with Francisella tularensis. These bacteria are some of the most virulent pathogens known, requiring inoculation or inhalation with as few as 10 organisms to cause disease. Infections can be acquired in a variety of ways: bites from an arthropod vector, skin lesions, ingestion of contaminated food or water, or inhalation of aerosolized particles. Its low infectious dose and relative ease of airborne transmission led to the development of F. tularensis as a biological weapon by several national weapons programs. Because it presents a serious threat to national security, the Centers for Disease Control (CDC) classify F. tularensis as a category A select agent. Despite these criteria, there is no approved tularemia vaccine. The goal of the research described herein is to construct a rationally attenuated living vaccine against tularemia. We will build upon our previous discovery of genes required for virulence in F. tularemia, mutant derivatives of which provide protection against wild-type challenge in mice. These genes will be deleted, individually and in combinations, in the most virulent subspecies of the organism, resulting in attenuated, non-reverting, defined strains that do not specify antibiotic resistance. After confirming that these strains protect mice against wild-type challenge, we propose to test the vaccines in a non-human primate model, a necessary step to achieving FDA approval. Tularemia is the severe and sometimes fatal disease caused by infection with Francisella tularensis. Its low infectious dose and relative ease of airborne transmission led to the development of F. tularensis as a biological weapon by several national weapons programs. As there is no approved tularemia vaccine, we seek to develop a live attenuated tularemia vaccine. [unreadable] [unreadable] [unreadable]